Primary brain tumours, such as gliomas, glioblastomas, meningiomas and pituitary adenomas, start from the neuroepithelium, ganglion cells, meninges, nerve sheaths, general nervous supporting tissue or neuroglia and pituitary or ectopic intracranial tissues (germ cell tumours or deformity tumours), and their causes are considered in particular to lie in genetic and hormonal factors, oncogenic viruses, and exogenous carcinogens. They are actual tumours of the central nervous system (CNS) localised in the brain, of varying differentiation and comprising various sub-types, such as: astrocytic tumours, oligodendrogliomas, mixed gliomas (oligoastrocytomas), ependymomas, tumours of the plexus choroideus, retinoblastomas, etc.
The WHO degree of classification is based on dignity: grade I (non-malignant, benign), grade II (semi-benign; postoperative life expectancy 3-5 years), grade III (semi-malignant; postoperative life expectancy 2-3 years), grade IV (malignant; postoperative life expectancy 6-16 months); and frequency: proportion of total or primary brain tumours in all tumour diseases: 7-9% (Kleihues, P., Louis, D. N., Scheithauer, B. W., Rorke, L. B., Reifenberger, G., Burger, P. C., and Cavenee, W. K. (2002) The WHO classification of tumors of the nervous system. J. Neuropathol. Exp. Neurol. 3, 215-225).
A glioma is characterised histologically in (giant cell) (oligo)astrocytoma, oligodendroglioma, mixed gliomas, glioblastoma, and is differentiated depending on growth, such as isomorphic, anaplastic, pilocytic, etc. Sub-groups of gliomas can also be predicted on the basis of the loss of heterozygosity (Smith, J. S., and Jenkins, R. B. (2000) Genetic alterations in adult diffuse glioma: occurrence, significance, and prognostic implications. Front Biosci. 5, 213-231), which lead to a loss of tumour suppressor genes (Tews, B., Felsberg, J., Hartmann, C., Kunitz, A., Hahn, M., Toedt, G., Neben, K., Hummerich, L., von Deimling, A., Reifenberger, G., and Lichter, P. (2006) Identification of novel oligodendroglioma-associated candidate tumor suppressor genes in 1p36 and 19q13 using microarray-based expression profiling. Int J Cancer. 119, 792-800).
In particular, glioblastomas (GBM) are one of the most aggressive brain tumours. The median survival of GBM patients, even with the best therapy preconditions, is only approximately 12-15 months. Natural killer (NK) cells as part of the natural immune system play a key role in the destruction of cancer cells. GBM cells develop strategies for avoiding this killing by down-regulating proteins that are necessary for interaction with NK cells, these being known as danger/stranger protein major histocompatibility complex (MHC), MHC class I polypeptide-related sequence (MIC)-A and -B, or UL16 binding proteins (ULBP) 1, 2, 3, more specifically by means of TGF-β mediated immunosuppression. To this end the overexpression of TGF-β is an essential feature of GBM, and high concentrations of TGF-β can be detected in glioma patients in the cerebrospinal fluid, which is correlated with the growth of the tumour (Kjellman C, Olofsson S P, Hansson O et al: Expression of TGF-beta isoforms, TGF-beta receptors, and SMAD molecules at different stages of human glioma. Int J Cancer 2000; 89: 251-258). In addition, TGF-β is responsible for the down-regulation of MHC expression, increasing the differentiation of naïve cells in T-reg cells, blocking dendritic cell maturation, and inducing cell death of K and T-cells (Eisele G, Wischhusen J, Mittelbronn M et al.: TGF-beta and metalloproteinases differentially suppress NKG2D ligand surface expression on malignant glioma cells. Brain 2006; 129: 2416-2425, Platten M, Wick W, Weller M: Malignant glioma biology: role for TGF-beta in growth, motility, angiogenesis, and immune escape. Microsc Res Tech 2001; 52: 401-410).
There is thus a great need to provide medicaments for the treatment and prophylaxis of brain tumours, in particular primary brain tumours, such as gliomas, glioblastomas, meningiomas and pituitary adenomas.
Mistletoe extracts have been used therapeutically for hundreds of years. In particular in cancer therapy, mistletoe preparations have been used with varying levels of success (Bocci V 1993 J Biol Regulators and Homeostatic Agents 7(1): 1-6; Gabius H-J, Gabius S, Joshi S S et al. 1993 Planta Med 60: 2-7; Gabius H-J & Gabius S 1994 PZ 139: 9-16; Ganguly C & Das S 1994 Chemotherapy 40: 272-278, Hajto T, Hostanska K, Gabius H_J 1989 Cancer Res 49: 4803-4808, Hajto T, Hostanska K, Frei K et al. 1990 Cancer Res. 50: 3322-3326). It has been found that the therapeutic effects are conveyed in particular by what are known as mistletoe lectins (viscumins, Viscum album Agglutinine, VAA). Mistletoe lectin, besides a cytotoxic effect, also brings about non-specific immunostimulation, the positive effects of which are used for therapy in tumour patients. Various studies with mistletoe lectin in vitro (Hajto et al., 1990 (supra); Mannel D N, Becker H, Gundt A et al. 1991 Cancer Immunol Immunother 33: 177-182; Beuth J, Ko K L, Tunggal L et al. 1993 Drug Res 43: 166-169) and in vivo (Hajto T 1986 Oncology 43 suppl 1: 51-65; Hajto et al., 1989 (supra), Beuth J, Ko H L, Gabius H-J et al. 1991 In Vivo 5: 29-32; Beuth J, Ko H L, Gabius H-J et al. 1992 J Clin Invest 70: 658-661), and clinical studies (Beuth et al., 1992 (supra)) have demonstrated an increased release of inflammatory cytokines (TNF-alpha, IL-1, IL-6) and an activation of cellular components of the immune system (TH-cells, NK-cells, B- and T-lymphocytes) (Braedel-Ruoff S: Immunomodulatory effects of Viscum album extracts on natural killer cells: review of clinical trials. Forsch Komplementmed 2010; 17: 63-73, Gren A: Effects of Iscador preparations on the reactivity of mouse immune system. Neuro Endocrinol Lett 2009; 30: 530-534, Lee C H, Kim J K, Kim H Y et al.: Immunomodulating effects of Korean mistletoe lectin in vitro and in vivo. Int Immunopharmacol 2009; 9: 1555-1561, Nikolai G, Friedl P, Werner M et al: Effect of a mistletoe extract (Iscador QuFrF) on viability and migratory behavior of human peripheral CD4+ and CD8+ T lymphocytes in three-dimensional collagen lattices. In Vitro Cell Dev Biol Anim 1997; 33: 710-716).
By analysis of the mistletoe extract, it has been possible thus far to identify three mistletoe lectins (ML-I, ML-II, ML-III) having different molecular weights and sugar-binding specificities. It has been found that the immunostimulating effect of the mistletoe extract on can be attributed to ML-I. The ML-I lectin consists of two glycosylated A- and B-chains (MLA and MLB). The A-chain is responsible for enzymatic inactivation of ribosomes (Endo Y, Tsurugi K & Franz H 1988 FEBS Lett 231: 378-380), whereas the B-chain is involved in carbohydrate bonding. The two chains are linked to one another by disulphide bridges. The resultant mistletoe lectin monomers can clump together to form dimers, with formation of non-covalent bonds.
It is possible to produce the biologically active mistletoe lectin advantageously recombinantly. EP 0751221 describes the preparation of mistletoe lectin polypeptides in a pure state as a structurally homogenous substance, wherein, proceeding from the gene sequence of mistletoe lectin, recombinant, highly pure individual chains (A-chain, B-chain) are produced, which can be re-associated in vitro and thus provide a recombinant mistletoe lectin holoprotein, which is homogeneous in respect of its protein chemistry, enzymatically and structurally, also known as Aviscumine. According to EP 0751221 the recombinant mistletoe lectin polypeptide is suitable as holoprotein and as sub-chain and in the form of sub-fragments for therapeutic purposes and is included within the scope of the invention.
Document WO2012104355A1 also describes the antiviral effect of recombinant mistletoe lectins. WO2012136857A1 discloses the treatment of skin cancer, in particular of a malignant melanoma also in the form of a metastasising tumour by means of recombinant mistletoe lectins.
Previously, recombinant mistletoe lectins were used advantageously in the treatment of tumour diseases. The use of recombinant mistletoe lectins for the treatment of brain tumours however, in particular primary brain tumours, such as gliomas, glioblastomas, meningiomas and pituitary adenomas, is not described in the prior art.
In the prior art, Podlech et al (Podlech O, Harter P N, Mittelbronn M et al.: Fermented mistletoe extract as a multimodal antitumoral agent in gliomas. Evid Based Complement Alternat Med 2012: 501796) describes the use of ISCADOR—a fermentatively obtained mistletoe extract—as growth inhibitor for GBM and indicates the antitumoral suitability of ISCADOR for the treatment of GBM.
Lenartz (Lenartz et al., Immunoprotective Activity of the Galactoside-Specific lectin from mistletoe after Tumor Destructive Theraly in Glioma Patients, Anticancer Research 16: 3799-3802 (1996)) reports on a mistletoe extract (ML-1) for the treatment of glioma patients, wherein the mistletoe extracts have a specific glycosylation.
The plant-based mistletoe lectins described in the prior art—whether or not obtained from fermentatively or non-fermentatively produced mistletoe extracts—are inhomogeneous (Soler M H, Stoeva S, Schwamborn C et al. 1996 FEBS Letter 399: 153-157, Soler H S, Stoeva S, Voelter W 1998 Biochem Biophys Res Comm 246: 596-601) and differ from one another non-uniformly in respect of their effect (EP 1051495 B1), and are not effective per se as active substance or as immunomodulator. Thus, the mistletoe lectin obtained from Korean mistletoe (Viscum album colaratum) for example should be assigned to the RIP II proteins, but has significant structural differences in the structure and conformation compared to the recombinant mistletoe lectin discussed here (Kang T B, Song S K, Yoon T J et al. 2007 J Biochem Mol Biol 40(6): 959-965). It is particularly disadvantageous that no exact dose adjustment is possible and that mistletoe lectins obtained from plant-based fermentatively or non-fermentatively produced extracts comprise impurities. Furthermore, the mistletoe lectins obtained from plant extracts—produced fermentatively or non-fermentatively—present differences in the glycosylation which influence the efficacy (in particular kinetics, etc.). Apart from this, the production of each new mistletoe extract batch yields a product that is not identical to the previous batch, with different contents of its ingredients including the glycosylated mistletoe lectins.